LOCUS       sequence-366023-        4897 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor with homologous arms for Rab11a to knock in 
            SFFV-T2A-Blasticidin resistance-PolyA cassette (for knock out).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4897)
  AUTHORS   Gemperle J, Harrison TS, Flett C, Adamson AD, Caswell PT
  TITLE     On demand expression control of endogenous genes with DExCon, 
            DExogron and LUXon reveals differential dynamics of Rab11 family 
            members.
  JOURNAL   Elife. 2022 Jun 16;11. pii: 76651. doi: 10.7554/eLife.76651.
  PUBMED    35708998
REFERENCE   2  (bases 1 to 4897)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Elife."; 
            date: "2022-06-16"; volume: "11. pii: 76651. doi"; pages: " 
            10.7554/eLife.76651"
COMMENT     Sequence Label: pJET-63-SFFV-T2A-BlaR-polyA-R11a
FEATURES             Location/Qualifiers
     source          1..4897
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     28..45
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    165..186
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(201..206,207..224,225..231)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     promoter        672..690
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        1017..1424
                     /label=SFFV promoter
                     /note="spleen focus-forming virus long terminal repeat 
                     (LTR) promoter"
     CDS             1523..1576
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             1586..1984
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     primer_bind     complement(2016..2033)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2022..2246
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        complement(2637..2655)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2961..2979)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3047..3151
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3152..3220,3221..4012)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3370..3389)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4183..4771
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4672..4691
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
       61 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
      121 aggtttcccg actggaaagc aattggcagt gagcgcaacg caattaatgt gagttagctc
      181 actcattagg caccccaggc tttacacttt atgcttccgg ctcgtataat gtgtggaatt
      241 atgagcggat aataatttca cacaggaggt ttaaacttta aacatgtcaa aagagacgtc
      301 ttttgttaag aatgctgagg aacttgcaaa gcaaaaaatg gatgctatta accctgaact
      361 ttcttcaaaa tttaaatttt taataaaatt cctgtctcag tttcctgaag cttgctctaa
      421 acctcgttca aaaaaaatgc agaataaagt tggtcaagag gaacatattg aatatttagc
      481 tcgtagtttt catgagagtc gattgccaag aaaacccacg ccacctacaa cggttcctga
      541 tgaggtggtt agcatagttc ttaatataag ttttaatata cagcctgaaa atcttgagag
      601 aataaaagaa gaacatcgat tttccatggc agctgagaat attgtaggag atcttctaga
      661 aagatagctg ataatacgac tcactatagg gactatggta gctaggagtt ccaggactca
      721 gtttcccctt tgagcctcct ttagcgacta aagcttgaag ccccacgcat ctcgactctc
      781 gcgcacaccg cccttgttgg gctcaggggc ggggcgccgc ccccggaagt acttcccctt
      841 aaaggctggg gcctgccgga aatggcgcag cggcagggag gggctcttca cccagtccgg
      901 cagttgaagc tcggcgctcg ggttacccct gcagcgacgc cccctggtcc cacagatacc
      961 actgctgctc ccgccctttc gctcctcggc cgcgcaatgg gatcgggtgg gactaggtaa
     1021 cgccattttg caaggcatgg aaaaatacca aaccaagaat agagaagttc agatcaaggg
     1081 cgggtacatg aaaatagcta acgttgggcc aaacaggata tctgcggtga gcagtttcgg
     1141 ccccggcccg gggccaagaa cagatggtca ccgcagtttc ggccccggcc cgaggccaag
     1201 aacagatggt ccccagatat ggcccaaccc tcagcagttt cttaagaccc atcagatgtt
     1261 tccaggctcc cccaaggacc tgaaatgacc ctgcgcctta tttgaattaa ccaatcagcc
     1321 tgcttctcgc ttctgttcgc gcgcttctgc ttcccgagct ctataaaaga gctcacaacc
     1381 cctcactcgg cgcgccagtc ctccgacaga ctgagtcgcc cgggggggat ctggagctct
     1441 cgagaattct cacgcgtctg caggatatca agcttgcggt accgcgggcc cgggatccat
     1501 ggtgagcact agtggcagcg gagagggcag aggaagtctt ctaacatgcg gtgacgtgga
     1561 ggagaatccc ggccctaggc tcgagatggc caagcctttg tctcaagaag aatccaccct
     1621 cattgaaaga gcaacggcta caatcaacag catccccatc tctgaagact acagcgtcgc
     1681 cagcgcagct ctctctagcg acggccgcat cttcactggt gtcaatgtat atcattttac
     1741 tgggggacct tgtgcagaac tcgtggtgct gggcactgct gctgctgcgg cagctggcaa
     1801 cctgacttgt atcgtcgcga tcggaaatga gaacaggggc atcttgagcc cctgcggacg
     1861 gtgccgacag gtgcttctcg atctgcatcc tgggatcaaa gccatagtga aggacagtga
     1921 tggacagccg acggcagttg ggattcgtga attgctgccc tctggttatg tgtgggaggg
     1981 ctaatctaga gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca
     2041 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     2101 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
     2161 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
     2221 tgctggggat gcggtgggct ctatgggtcg acggaaccgg tgctggaagt ggtacacgcg
     2281 acgacgagta cgactacctc tttaaaggtg aggccatggg ctctcgcact ctacacagtc
     2341 ctcgttcggg gacccgggcc actcccggtg gaccctcgtg ccggccaccc ctgcactgat
     2401 ataggcctcc ctcagccctt cctttttgtg cggttccgtc tcctacccag ctcagcctct
     2461 tctcccccgc tcagacaggg gtccccatca catgccgctc tctgagcgac ctctccatag
     2521 gccttcgctg gcctcagagc ccctccctgc gtgtccttcc cctggcggac tgccttctcc
     2581 cacatcgtat cttgctgaaa aactcgagcc atccggaaga tctggcggcc gctctcccta
     2641 tagtgagtcg tattacgccg gatggatatg gtgttcaggc acaagtgtta aagcagttga
     2701 ttttattcac tatgatgaaa aaaacaatga atggaacctg ctccaagtta aaaatagaga
     2761 taataccgaa aactcatcga gtagtaagat tagagataat acaacaataa aaaaatggtt
     2821 tagaacttac tcacagcgtg atgctactaa ttgggacaat tttccagatg aagtatcatc
     2881 taagaattta aatgaagaag acttcagagc ttttgttaaa aattatttgg caaaaataat
     2941 ataattcggc tgcaggggcg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     3001 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     3061 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     3121 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     3181 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     3241 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     3301 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     3361 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     3421 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     3481 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     3541 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     3601 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     3661 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     3721 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     3781 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     3841 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     3901 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     3961 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     4021 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     4081 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     4141 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     4201 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     4261 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     4321 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4381 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4441 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4501 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4561 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4621 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     4681 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     4741 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     4801 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     4861 ggataaccgt attaccgcct ttgagtgagc tgatacc
//
